4.P.302 Novel elements located at −504 to −399 bp of the promoter region regulated the expression of the human macrophage scavenger receptor gene in murine macrophages

Liao, Hai-Sun; Kodama, Tatsuhiko; Doi, Takefumi; Emi, Mitsuru; Asaoka, Hiroshi; Itakura, Hiroshige; Matsumoto, Akiyo

doi:10.1016/s0021-9150(97)89827-9

Cited by 7 publications

(11 citation statements)

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“…21 Briefly, suspension cultures of 5ϫ10 6 cells were transfected with 3 g of reporter gene along with either 3 g PPAR␥/pcDNA3 or pcDNA3 by setting the capacitance at 975 F and voltage at 250 V. C2/2 cells were transfected with a series of the OPN reporter construct (0.1 g) along with either 1 g of PPAR␥/pcDNA3 or pcDNA3 as previously described. 20 After transfection, cells were exposed to PMA (100 ng/L) alone and in combination with Tro (10 mol/L).…”

Section: Transfection and Luciferase Assaysmentioning

confidence: 99%

PPARγ Ligand Inhibits Osteopontin Gene Expression Through Interference With Binding of Nuclear Factors to A/T-Rich Sequence in THP-1 Cells

Oyama

Akuzawa

Nagai

et al. 2002

Circulation Research

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Abstract-Peroxisome proliferator-activated receptor ␥ (PPAR␥) is a member of the nuclear receptor superfamily that acts as a key player in adipocyte differentiation, glucose metabolism, and macrophage differentiation. Osteopontin (OPN), a component of extracellular matrix, is elevated during neointimal formation in the vessel wall and is synthesized by macrophages in atherosclerotic plaques. In the present study, we investigated the molecular mechanisms regulating OPN gene expression by PPAR␥ in THP-1 cells, a cell line derived from human monocytic leukemia cells. Northern and Western blot analyses showed that exposure of THP-1 cells to PMA (phorbol 12-myristate 13-acetate) increases OPN mRNA and protein levels in a time-dependent manner. PMA-induced OPN expression was significantly decreased by troglitazone (Tro) and other PPAR␥ ligands. Transient transfection assays of the human OPN promoter/luciferase construct showed that PPAR␥ represses OPN promoter activity, and the PPAR␥-responsive region within the OPN promoter lies between Ϫ1000 and Ϫ970 relative to the transcription start site. Site-specific mutation analysis and electrophoretic mobility shift assays indicated that a homeobox-like A/T-rich sequence between Ϫ990 and Ϫ981, which

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Section: Transfection and Luciferase Assaysmentioning

confidence: 99%

PPARγ Ligand Inhibits Osteopontin Gene Expression Through Interference With Binding of Nuclear Factors to A/T-Rich Sequence in THP-1 Cells

Oyama

Akuzawa

Nagai

et al. 2002

Circulation Research

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Section: Assays For Msr Promoter Activitymentioning

confidence: 99%

“…31 THP-1 cells were cotransfected with luciferase constructs and the chloramphenicol acetyltransferase (CAT)-control plasmid (as an internal control) by electroporation. 31 In brief, THP-1 cells at 5ϫ10 5 /mL were plated 16 to 24 hours before transfection. Electroporation was then carried out at room temperature under 960 F and 350 V in 700 L of the reaction mixture containing 2ϫ10 7 cells, 30 g of the indicated luciferase construct, and 10 g of CAT-control plasmid DNA in a 0.4-cm electroporation cuvette (Bio-Rad).…”

Section: Assays For Msr Promoter Activitymentioning

confidence: 99%

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Transcriptional Inhibition by Interleukin-6 of the Class A Macrophage Scavenger Receptor in Macrophages Derived From Human Peripheral Monocytes and the THP-1 Monocytic Cell Line

Liao

Matsumoto

Itakura

et al. 1999

ATVB

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Abstract-Expression of the class A macrophage scavenger receptor (MSR) contributes to the uptake of modified low density lipoproteins (LDL) by macrophages and transformation of these cells into lipid-laden foam cells, which characterize atherosclerosis. Many environmental factors, in particular, proinflammatory cytokines and growth factors, can exert regulatory effects on MSR expression, whereas intracellular accumulation of cholesterol itself does not influence MSR levels to any considerable extent. In the present study, by using an in vitro model, we examined whether stimulation with interleukin-6 (IL-6), an immunoregulatory, multipotential cytokine, modulates the expression and activities of the MSR in macrophages. When treated with IL-6, macrophages derived from peripheral monocytes and phorbol 12-myristate 13-acetate (PMA)-differentiated THP-1 monocytic cells showed significantly reduced uptake and/or binding of the MSR ligand, acetylated LDL. This effect was paralleled by a reduction in the expression of MSR protein and mRNA. Analysis of MSR promoter activity in THP-1 cells transfected with an MSR promoter-reporter gene construct demonstrated decreased activity of the MSR promoter in IL-6 -treated THP-1 macrophages. Electrophoretic mobility gel shift assay also showed a reduction in the binding of a transcription factor to the MSR promoter AP-1/ets elements in IL-6 -treated cells. Thus, exposure to IL-6 may inhibit expression of the class A MSR in differentiated macrophages at transcriptional levels. This result suggests that this cytokine may modulate foam cell formation during atherogenesis.

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“…21 Briefly, an oligonucleotide (5'-AACGCAGGAATGTGTCATTTCCTTT-3') coding for the AP1/ets-like cis-element extending from 710 to +15 bp of the human MSR promoter labeled with g-32 P-ATP and T4 polynucleotide kinase was mixed with cell nuclear extracts. The binding reaction between the oligonucleotide and nuclear proteins was performed in 12.5 ml reaction mixtures containing 4 mM Tris-HCl (pH 7.9), 10 mM HEPES (pH 7.9), 1 mM DTT, 1 mM EDTA, 60 mM KCl, 0.2 mg/ml poly (dI-dC), and 10% glycerol with 6 mg cell extract protein.…”

Section: Gel Electrophoretic Mobility Shift Assay (Emsa)mentioning

confidence: 99%

De novo expression of the class-A macrophage scavenger receptor conferring resistance to apoptosis in differentiated human THP-1 monocytic cells

et al. 1999

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The class-A macrophage scavenger receptor (MSR) is a trimeric multifunctional protein expressed selectively in differentiated monomyeloid phagocytes which mediates uptake of chemically modified lipoproteins and bacterial products. This study investigated whether MSR plays a role in the regulation of apoptosis, a model of genetically programmed cell death. De novo expression of MSR occurred in human THP-1 monocytic cells differentiated with phorbol esters, which activated a nuclear transcription factor binding to the Ap1/ets-like domain of the MSR promoter. The phorbol ester-stimulated THP-1 cells also expressed increased levels of the pro-apoptotic gene products, caspase-3 and Fas ligand, but the cells exhibited no change in apoptosis. Global activation of GTP-binding proteins with fluoride anions triggered apoptosis of THP-1 cells in a time-and concentration-dependent manner, demonstrated by nuclear shrinkage and fragmentation and internucleosomal DNA fragmentation. However, the MSR-expressing THP-1 macrophage-like cells showed a significant reduction in apoptosis compared to undifferentiated control THP-1 cells, which produce MSR at undetectable levels. Fluoride stimulation also triggered apoptosis of human Jurkat T cells. Stimulation with phorbol ester made no difference in apoptosis between treated and untreated Jurkat cells. Finally, Chinese hamster ovary (CHO) cells overexpressing the class-A MSR type I by cDNA transfection showed markedly increased resistance to Gprotein-coupled apoptosis. Thus, de novo expression of MSR associated with monocyte maturation into macrophages appears to confer the resistance of macrophages to apoptotic stimulation by G-protein activation.

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4.P.302 Novel elements located at −504 to −399 bp of the promoter region regulated the expression of the human macrophage scavenger receptor gene in murine macrophages

Cited by 7 publications

References 0 publications

PPARγ Ligand Inhibits Osteopontin Gene Expression Through Interference With Binding of Nuclear Factors to A/T-Rich Sequence in THP-1 Cells

PPARγ Ligand Inhibits Osteopontin Gene Expression Through Interference With Binding of Nuclear Factors to A/T-Rich Sequence in THP-1 Cells

Transcriptional Inhibition by Interleukin-6 of the Class A Macrophage Scavenger Receptor in Macrophages Derived From Human Peripheral Monocytes and the THP-1 Monocytic Cell Line

De novo expression of the class-A macrophage scavenger receptor conferring resistance to apoptosis in differentiated human THP-1 monocytic cells

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